Age is one of the major risk factors for development of the liver cancer, which is the fifth most common cancer and the third most common cause of cancer related death in the world. Despite significant advances in cancer research, little progress has been made in early detection and treatment of liver cancer leading to poor prognosis. Over the past 15 years, my laboratory has pursued the studies to determine the key signaling pathways that are changed by aging and that make liver cells more susceptible to development of liver cancer. One of these pathways is the hyper-phosphorylation of a transcription factor, CCAAT/Enhancer Binding protein alpha, C/EBPa, at S193. We have successfully generated C/EBPa-S193D knockin mice (S193D) which mimic the age-associated ph-S193 isoform of C/EBPa and found that liver proliferation after partial hepatectomy is completely inhibited in S193D mice. To our surprise, we found that the development of liver tumors in S193D mice occurs much faster than in WT mice. Furthermore, we found that gankyrin specifically interacts with and degrades the S193D and ph-S193 isoforms of C/EBPa. The main hypothesis of the application is that the activation of gankyrin and following degradation of C/EBPa is a key regulatory pathway in development of liver cancer. Our recent studies show that farnesoid X receptor (FXR) and its down-stream target SHP suppress gankyrin. The deletion of FXR/SHP leads to activation of gankyrin resulting in the degradation of C/EBPa and the formation of spontaneous liver tumors at age between 12-17 months. Specific Aim 1 will address mechanisms by which FXR/SHP pathway represses gankyrin. In this aim, we plan to accomplish the following: a) Investigate if single FXR and SHP KO mice activate gankyrin, b) Examine the activation of gankyrin in newborn, 2 mo, 6 mo and 12 mo old FXR/SHP DKO mice in order to determine the age at which gankyrin is activated, c) Identify FXR-dependent transcription factors that are involved in the activation of gankyrin, and d) Verify the activation of gankyrin- C/EBPa pathway by measuring gankyrin in human HCC samples obtained from patients of different ages. Specific Aim 2 will examine if the activation of FXR inhibits liver tumor. We will activate FXR by FXR-specific ligand GW4064 and examine if this activation will reduce the incidence/number of liver tumors in DEN- mediated carcinogenesis. Specific Aim 3 will investigate if the inhibition of gankyrin-C/EBP1 pathway will prevent liver tumor formation. We plan to use the following approaches: a) Inhibit the expression of gankyrin by siRNA in both WT DEN-treated mice and FXR/SHP mice and examine if this inhibition will reduce the incidence/number of liver tumors, and b) Develop approaches to block gankyrin binding to C/EBP1. Thus, the proposed studies will provide novel gankyrin-based approaches for the prevention of liver cancer. PUBLIC HEALTH RELEVANCE: This project investigates molecular basis for the development of cancer in the liver. Despite significant advances in cancer research, there is an urgent need to develop new approaches for the early detection and effective management of liver cancer. We will determine mechanisms by which liver cancer activates gankyrin and degrades tumor suppressor ph-S193 isoform of C/EBP1 with the goal to provide basement for the generation of therapeutic approaches to prevent liver cancer.